Recent advances have been made in mutating or inactivating genes in murine embryonic stem (ES) cells via homologous DNA recombination with exogenously added DNA. The ES cells have the ability to differentiate into somatic tissue when injected into a 3.5 day blastocysts and can even contribute to the germline. Therefore, a tissue culture generated mutation can eventually give rise to new mouse strains with the desired mutation. This powerful new technology is well suited to help analyze the developing immune system. The vertebrate immune system has been extensively studied at the level of cell-cell interactions and the development of many of the cell-types involved in an immunological response. T cells comprise a major portion of the cellular response to infectious agents and can be broadly grouped into CD8 + cytotoxic T cells (CTL), CD4 helper T (Th) cells and the recently discovered gamma-delta T cells. In the thymus CTL and Th are positively selected to recognize MHC molecules as self but are deleted if too strong an interaction (autoreactivity) with MHC occurs. In this process T cells are "educated" in self/non-self discrimination. In the periphery, cells encounter and process foreign antigen and present peptide fragments of the antigen in association with MHC. T cells recognize the MHC-antigen complex as foreign and mount a response to eliminate the infectious agent. This proposal describes the methodology to generate MHC class II negative strains of mice by using homologous recombination in ES cells, The elimination of the I-A(b) molecule by inactivation of the I-A(beta) gene will address the issue of the role of that molecule in helper T cell development, the potential importance of I-A in CTL development, and the ability of mice lacking these genes to clear infectious agents.